Method for production of corosolic acid in suspension culture of plant cells

Abstract

The present invention relates to a method of producing corosolic acid by using plant cells that produce corosolic acid. More particularly, the present invention relates to a method of producing corosolic acid by using plant cell suspension cultures comprising the steps of: inducing a callus from a tissue of a plant producing corosolic acid; preparing a cell line capable of being cultured in liquid culture medium from the induced callus; culturing the cell line in a suspension culture; and isolating corosolic acid from the culture solution. The present invention has advantage of maximizing productivity by utilizing two-stage culture, treatment with inducing agent, and high cell-density culture in the suspension culture of plant cells producing corosolic acid.

1. A method of producing corosolic acid by using plant cell suspension culture comprising the steps of:
 (a) inducing a callus from a tissue of plant producing corosolic acid; 
 (b) preparing a suspension-cultured cell line capable of being cultured in liquid culture medium from the induced callus; 
 (c) culturing the prepared suspension-cultured cell line in a suspension culture; 
 (c-1) culturing by transferring the cell line to a medium containing an elicitor: and 
 (d) isolating corosolic acid from the culture solution obtained in the suspension culture step (c-1), wherein the elicitor is selected from the group consisting of biological elicitors and non-biological elicitors. 
 
     
     
       2. The method of  claim 1 , wherein the plant producing corosolic acid is selected from the group consisting of  Eriobotrya japonica, Lagerstroemla speciosa, Temstroemia gymnanthera, Crataegus pinnatifida , and  Tiarella polyphylla.    
     
     
       3. The method of  claim 1 , wherein the suspension-cultured cell line is a cell line of  Eriobotrya japonica  Lindl. SYG-2 (accession no. KCTC 10822BP), or a cell line of  Lagerstromia speciosa  SYG-3 (accession no. KCTC 10823BP). 
     
     
       4. The method of  claim 1 , wherein in step (a), the callus is induced by culturing the plant tissue on Linsmaler & Skoog medium, or Murashige & Skoog medium. 
     
     
       5. The method of  claim 1 , wherein the medium further comprises one or more selected from the group consisting of α-naphtalene acetic acid and 6-benzylaminopurine. 
     
     
       6. The method of  claim 1 , wherein in step (c), the cell line is in Linsmaler & Skoog medium, or Gamborg's B5 medium. 
     
     
       7. The method of  claim 1 , wherein the medium of step (c-1) is Gamborg's B5 medium or modified Gamborg's B5 medium. 
     
     
       8. The method of  claim 1 , wherein the elicitor is treated one or more times. 
     
     
       9. The method of  claim 1 , wherein the non-biological elicitor is selected from the group consisting of silver nitrate, sodium butyrate, and methyl jasmonate. 
     
     
       10. The method of  claim 9 , wherein silver nitrate is treated at a concentration of 0.01 to 500 μM. 
     
     
       11. The method of  claim 9 , wherein sodium butyrate is treated at a concentration of 0.01 to 500 mM. 
     
     
       12. The method of  claim 9 , wherein methyl jasmonate is treated at a concentration of 0.1 μM to 50 mM. 
     
     
       13. The method of  claim 1  wherein the biological elicitor is selected from the group consisting of an extract of fungi, and extract of bacteria, and extract of yeast, chitosan, lichenan, glucomannan, pleuran, glucan, carboxylmethylglucan, sulfoethylglucan, hydroxymethylglucan, mannan, xylan, mannobiose, mannotriose, mannopentaose, mannotetraose, cellulysin, Multifect XL, Multifect CL, resinase, pilpxyme, SP 431, pectinal, rapidase, and chitinase. 
     
     
       14. The method of  claim 1  wherein the initial concentration of inoculated cells is 1 to 20 g/L DW (cell dry weight). 
     
     
       15. The method of  claim 1  wherein the medium comprises a carbon source in an amount of 1 to 20%(w/v). 
     
     
       16. A method of suspension culture of  Eriobotrya japonica  or  Lagerstromia speciosa  comprising the steps of:
 (a) inducing a callus from  Eribotrya japonica  or  Lagerstromia speciosa;    
 (b) preparing a cell line capable of being cultured in liquid culture medium from the induced callus; 
 (c) culturing the cell line in a suspension culture; and 
 (d) culturing by transferring the cell line to a medium containing an elicitor. 
 
     
     
       17. The method of  claim 16 , wherein the cell line is a cell line of  Eriobotrya japonica  Lindl. SYG-2 (accession no. KCTC 10822BP), or a cell line of  Lagerstromia speciosa  SYG-3 (accession no. KCTC 10823BP).